REPETITIVE DNA

DNA may be categorised as :-

Single copy DNA sequences (60% of total)

• Present in single or low copy numbers.
• Includes coding sequence for structural genes (up to 1400 bp in size), which account for 3% of the genome.
• The remainder is intronic sequence or spacer DNA.

Moderately repetitive DNA sequences (30% of total)

• Present at between 10 - 10⁵ copies per genome. Found throughout the euchromatin.
• Average 300bp in size
• May be classed as :-
  

  a) microsatellites / minisatellites (VNTR, DNA 'fingerprints)
  b) dispersed-repetitive DNA, mainly transposable elements (LINES/ SINES)

   

• Also includes 'redundant' genes for histones, and ribosomal RNA and proteins, (gene-products present in cell in large numbers).

   

• Many moderately-repetitive sequences may be involved in regulation of gene expression. This is supported by their interspersion with single-copy sequences and location adjacent to structural genes.

Highly repetitive DNA sequences (satellite DNA) (10% of total)

• Present at >10⁶ copies per genome
• Occurs as variable length motifs (5-100 bp), in long tracts of up to 100 Mb
• Most is located in heterochromatic regions around the centromere / telomere.
• Postulated functions include structural or organisational roles, role in chromosome pairing, involvement in cross-over or recombination, junk.

  eg. alpha-satellite DNA
  This is a highly repetitive sequence, each centromere contains a tandem array of alpha-satellite repeats that extend for millions of base pairs and are arranged in a hierarchy of higher order repeats. These vary between 100-5000 on different chromosomes (0.2-10Mb). Some contain 17bp binding sites for the centromere-specific DNA binding protein CENP-B. They have been recently cloned and used to construct artificial human chromosomes.
  

Much of the genome consists of moderately-repetitive sequences interspersed with single copy sequences.

Evolution of repetitive DNA
Microsatellites tend to be highly polymorphic, suggesting a 'stepwise mutation' model in which most variation is introduced by replication slippage, changing the array length by only one or two repeats at a time, but also with occasional larger 'jumps' in size at much lower frequency.

Minisatellites, evolve more readily by larger-scale mechanisms such as unequal exchange. For all classes there appears to be a general bias towards increase in array length through evolutionary time.

Highly repetitive DNA tends to accumulate only in regions of low recombination such as centromeres and telomeres, where recombination is suppressed, while repeats occurring in euchromatin are much more susceptible to crossing-over and tend to be more variable in copy number relative to their array length.

Transposable elements (mobile genetic elements)
Much of moderately-repeated DNA consists of transposable elements. The two major families, the long and short interspersed nucleotide elements (LINEs and SINEs), are represented in humans mainly by L1 and Alu elements respectively. Both types of element are considered to be retrotransposable (ie. can replicate via an RNA copy reinserted as DNA by reverse transcription) and they have significant roles in genomic function and evolution.

The majority of inserted elements are truncated and often rearranged relative to full-length elements;

The major transposable elements in humans are :-
LINES (long interspersed elements) and SINES (short interspersed elements)
The most common examples in humans are L1 and Alu elements which are thought to have arisen by retrotransposition.

The most common and best characterised LINE is L1

• Repeated approx. 50,000x in the human genome (0.5% of total)
• Only 3000 of these are full length; the remainder are truncated, mostly at the 5' end.
• Complete element is 6kb in size and contains two open reading frames, one of which encodes a reverse transcriptase.
• AT-rich region is located near the 3' end of the element,
• Element is flanked by two short direct repeats

The main type of SINE is the Alu family
(So named as they usually contain a target for the restriction enzyme Alu I).

• 5 x 10⁵ - 10⁶ copies in the haploid genome, with an average of one repeat every 4kb (1 - 10 % total)
• Not found within coding regions but often present in the transcription unit, within introns and occasionally in non-translated regions of the mRNA.

   

• All contain 290bp consensus sequence which consist of two tandem repeats of a 130bp sequence, one of which has a 32bp deletion.

   

• Elements are flanked by direct repeats
• Each repeat unit has an AT-rich region that suggests a poly A tail
• 5' end resembles a pol III promoter region.

   

• The origin of Alu can be traced to the terminal portions of the signal recognition particle 7SL RNA, hence it is probably a pseudogene formed by loss of the 7SL RNA central fragment.

LINEs and SINEs both have a poly(A) tail which may act as a template for reverse transcription from nicks made at the site of insertion in the host DNA by a LINE-encoded endonuclease. In contrast, Alu is transcribed but not translated. For transposition to proceed it is thought that the Alu RNA needs to 'hijack' the endonuclease/reverse transcriptase protein encoded by L1 to complete its insertion.

How can they cause disease?

Although TEs do not contribute to the phenotype, they can affect it,and retrotransposal integrations of Alu and L1 sequences into biologically important genes appear to play significant roles in some human diseases. However, although a large number of TEs are transcriptionally active, only a small subset (<0.01%) are able to transpose ie. capable of causing mutations.

Evidence for insertional mutagenesis by SINEs and LINEs in mammals is becoming more abundant, with L1 in particular having been demonstrated in cases of haemophilia, DMD, and sporadic breast and colon cancer. Integrations have been observed in oncogenes and in tumor suppressor genes which may participate in carcinogenesis by altering gene activity. The exact mechanism of these events is unclear

Recent evidence has demonstrated the integration of Alu sequences into control regions, where they can bind regulatory proteins and modulate transcription.

Unequal crossing-over between repetitive elements may be the cause of gene duplication, (from which gene families arose).

References

Charlesworth B, Sniegowski P, Stephan W (1994) The evolutionary dynamics of repetitive DNA in eukaryotes. Nature 371, 215-220

Epplen C, Santos EJ, Maueler W, van Helden P, Epplen JT (1997) On simple repetitive DNA sequences and complex diseases. Electrophoresis 18, 1577-85

Miki Y (1998) Retrotransposal integration of mobile genetic elements in human diseases, J Human Genetics 43 (2) 77-84

Mighell AJ, Markam AF, Robinson PA (1997) Alu sequences.
FEBS Letters 417 (1) 1-5

Peter Sudbery. Human Molecular Genetics.
(Cell and Molecular Biology in Action series) 1998